Reading& Problems: LNC p. 200-206; p. 238 prob. 8, 11, 13; Segel, p.319, prob. 1,2,3
Rate of production of product is the velocity V = d[P]/dt
- Initial rate - we are in the initial phase of the reaction when the reaction has proceeded to a small enough extent that the [S] does not change significantly.
- Steady state - the [ES] rapidly reaches a steady state that does not change over the time being analyzed.
D. Plotting enzyme data (here is a link to download the enzyme plots used in class)
- [P] vs. time
- V versus [S]
A. Lineweaver-Burk double reciprocal plot: 1/V vs 1/[S] and determination of Km (X-intercept is -1/Km) and Vmax (Y-intercept is 1/Vmax), slope is Km/Vmax
- Km - the [S] that leads to half maximal velocity, a measure of the affinity of the enzyme for the substrate. Range 0.4 µM - 10 mM. Lower Km means higher affinity.
- Vmax - not really a constant, =kcat[Et] and [Et] can vary. It is only a constant at a constant enzyme concentration.
- Kcat - how many reactions one enzyme can perform per unit time. In inverse time units. In sec-1 (per second) called "turnover number". Range from less than 0.5 up to 6 x 105. Higher Kcat means faster reaction.
- Best measure of enzyme efficiency is Kcat/Km. Bigger number is more efficient enzyme.
Some take home information:
Lineweaver-Burk double reciprocal plot: 1/V vs 1/[S] can be used to determine Km (X-intercept is -1/Km) and Vmax (Y-intercept is 1/Vmax)
Charles S. Gasser (Department of Molecular & Cellular Biology; UC Davis)